HepG2 cells (5 × 105 cells) in a six-well plate were incubated (37°C, 5% CO2) with 100 μg/mL extracts or sorafenib or DMSO vehicle (<0.5%) (as control) media for 24 h. Following trypsinization, cells were washed and centrifuged at 2000 ×g for 10 min and the pellet was resuspended in 0.5 mL PBS. Fixation was completed by adding 1.2 mL of 70% cold ethanol for 2 h. The fixed cells were washed with PBS and centrifuged at 2000 ×g for 10 min. After suspending cells in 0.3 mL PBS, 8 μL of DNAase free RNAse (10 mg/mL) was added and incubated for 1 h. After adding 15 μL of propidium iodide (0.5 mg/mL), cells were incubated at 4°C for 30 min. The cells were analyzed for cell cycle using flow cytometer FACS calibur (Beckman Coulter, Fullerton, CA, USA) with an excitation wavelength of 488 nm and emission at 670 nm. DNA content was determined by ModFit software (Verity Software House, Topsham, ME), which provided histograms to evaluate cell cycle distribution.
Flow cytometer facs calibur
Manufactured by Beckman Coulter
Sourced in United States
The FACS Calibur is a flow cytometer designed for multi-parameter analysis of cells and other particles. It utilizes laser-based technology to detect and measure various physical and fluorescent characteristics of individual cells or particles as they flow through the instrument. The FACS Calibur is capable of analyzing multiple parameters simultaneously, providing detailed information about the sample under investigation.
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Lab products found in correlation
2 protocols using flow cytometer facs calibur
1
Cell Cycle Analysis of HepG2 Cells
2
Cell Cycle Analysis of Fatty Acid Esters
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HepG2 cells were plated at 5×105 cells per ml in a six-well plate. After 24 h incubation (37°C, 5% CO2), the cells were treated with 100 µM fatty acid esters of phloridzin, phloridzin, phloretin, sorafenib or DMSO (<0.5%) control prepared in media and incubated for additional 24 h. Following trypsinization, cells were washed and centrifuged at 2000×g for 10 min and the pellet re-suspended in 0.5 mL PBS. Fixation was completed by adding 1.2 mL of 70% cold ethanol for 2 h. The fixed cells were washed with PBS and centrifuged at 2000×g for 10 min. After suspending cells in 0.3 mL PBS, 8 µL of DNAase free RNAse (10 mg/mL) was added and incubated for 1 h. After adding, 15 µL of propidium iodide (0.5 mg/mL), cells were incubated in 4°C for 30 min. The cells were analyzed for cell cycle using flow cytometer FACS calibur (Beckman Coulter, Fullerton, CA, USA) with an excitation wavelength of 488 nm and emission at 670 nm. DNA content was determined by ModFit software (Verity Software House, Topsham, ME, USA), which provided histograms to evaluate cell cycle distribution.
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